Project Summary ? Abstract The perception of odors begins in the olfactory epithelium when odorant ligands bind to molecular receptors expressed on the cilia of the olfactory sensory neurons, each of which expresses only 1 of ~1200 candidate receptors. As the sensory neuron axons exit the epithelium they progress over the surface of the olfactory bulb and all of the axons coming from neurons expressing the same odorant receptor converge into only 2/3 glomeruli/olfactory bulb. However, the convergence primary afferent axons within the glomerulus and the discrete local circuitry modulating afferent input remains uncertain Despite a long history of interest, we remain woefully ignorant of the most fundamental features of cellular and synaptic organization within glomeruli: What is the distribution of the dendritic processes of projection neurons versus interneurons within the glomerulus? What is the 3-dimensional topography of synapses along dendrites and in the core versus the periphery of the glomerulus? What is the nature and composition of non-synaptic interactions among dendrites? With the recent development of Serial Block-Face Scanning Electron Microscopy (sbSEM) the ability to serially reconstruct glomeruli at the ultrastructural level, to establish the glomerular connectome, these and other questions are within our grasp. To begin addressing the utility of sbSEM in understanding olfactory bulb circuitry we begin with with the following: 1) The hypothesis that the dendrodendritic synapses between glomerular interneurons and projection neurons are reciprocal. The dendritic Gray Type I and II synapses in glomeruli appear unipolar and isolated. They are presumed to be reciprocal based on physiology, but if there is reciprocity, it is not known if the ratio is 1:1 or unequal. Moreover, it is not clear if they mediate self- or lateral inhibition. 2) The hypothesis that axodendritic (excitatory) and dendrodendritic (inhibitory) synapses differentially localize to the distal versus the proximal dendritic segments of projection neurons. Gate-keeping inhibitory synapses are often found at dendritic branch points, but whether that holds true in glomeruli must be empirically determined using comprehensive 3D dendritic reconstructions. 3) The hypothesis that single OSN axon establish divergent synaptic connections with dendrites from different neurons. Does a single axon repeatedly contact the same dendrite, or is it divergent? While the question seems simple, the answer will provide new insight into the roles of feed- forward and feed-back processing. 4) The location(s) of presynaptic inhibition on OSN axons. Electrophysiological analyses support presynaptic inhibition by showing, for example, that GABAB agonists decrease Ca++ influx into the axon terminals. However, the site of synaptic apposition is not known. Using sbSEM to address these questions will resolve fundamental controversies regarding processing of odors within the glomerulus and provide the foundation for further studies of development and plasticity.